The ability to introduce foreign molecules into living cells and human tissues has significant implications for various applications in biological research and medicine. Numerous methods including virus-mediated, chemical, physical, and optical approaches have been developed in order to deliver exogenous molecules into the cells. Currently, virus-mediated approach is the most efficient way of gene delivery and expression; however, besides being capable only for delivery of nucleic acids, there are considerable risks accompanied with viral delivery systems, including toxicity, chromosomal integration and immunogenicity. Viral approaches are, hence, less than ideal for many clinical and research applications, which require minimal post transplantation risks, such as gene therapies and studies of cellular reprogramming or lineage conversions.
Consequently, non-viral molecular delivery techniques have begun to gain more attention as inevitable alternatives. Among these techniques, electroporation is considered as an effective and powerful technique because of its ability to introduce countless types of molecules into target cells both in vitro and in vivo without need for potentially cell-damaging chemical reagents or viruses. Electroporation utilizes short high-voltage pulses to transiently and reversibly create pores on cell membrane through which molecular probes of interest can be delivered into the cytosol. However, conventional electroporation techniques using cuvettes or micro-capillaries rely on bulk stochastic molecule delivery processes, making those systems ill-suited for applications requiring precisely and individually controlled transferred molecular doses. Moreover, it is difficult to obtain practical efficiency and viability for samples with large heterogeneity in cell diameter since the electric field strength required to transiently disrupt cellular membrane strongly correlate with the cell size.
Recent advances in microfluidics facilitated development of microscale electroporation techniques, allowing the single-cell level electroporation and dosage control of transferred molecules with enhanced cell viability. Despite their noteworthy improvement in throughput and molecular delivery efficiency with enhanced viability, a single-directional flow-through scheme under which most of current microfluidic electroporation systems operate renders current microfluidic electroporators do not provide for multiple different molecules and also lack good dosage control.